Differential cytokine gene expression profiles in the three pathological forms of sheep paratuberculosis.

Smeed JA, Watkins CA, Rhind SM, Hopkins J - BMC Vet. Res. (2007)

Bottom Line:
No changes were seen in IL-1 alpha in any sheep ileum tissues.Asymptomatic animals are IS900+ with normal histology but have significantly decreased levels of IL-18 and increased levels TNFalpha.Asymptomatic animals are infected and show no pathology but can be distinguished, in terms of cytokine expression pattern, from uninfected controls.

Background: Johne's disease is a chronic inflammatory disease of the gut caused by infection with Mycobacterium avium subspecies paratuberculosis (MAP). Symptoms include wasting, diarrhoea, loss of condition and eventual death. Three forms of Johne's disease have been described in sheep - paucibacillary, multibacillary and asymptomatic. The paucibacillary form is characterized by an inflammatory, Th1-type immune response. The multibacillary form of the disease, which disseminates the infection, is characterized by macrophage infiltration mediated by a Th2-type immune response, and asymptomatic animals have no clinical symptoms or pathology but are infected with MAP. What determines these three forms of the disease is unknown. To further understand these differences, we used real-time RT-PCR to compare the expression of thirteen cytokine and cytokine-related genes in ileal tissue from sheep with the three forms of the disease.

Results: Three pathological forms of sheep paratuberculosis were defined on the basis of histopathology, cytochemistry (Zeihl-Neelsen) and IS900 PCR. Paucibacillary lesions have largely T cell and eosinophil infiltration and are ZN negative; multibacillary lesions have macrophage infiltration and large numbers of acid-fast bacteria. The pauci- and multibacillary forms are linked to the differential expression of IFN gamma and IL-10 respectively. In addition the increased levels of the proinflammatory cytokines (IL-1 beta and TNFalpha), IL-8, IL-18 and TRAF-1 in both diseased forms is indicative of persistent inflammatory lesions. No changes were seen in IL-1 alpha in any sheep ileum tissues. Asymptomatic animals are IS900+ with normal histology but have significantly decreased levels of IL-18 and increased levels TNFalpha.

Conclusion: We have quantified the expression levels of thirteen cytokine and cytokine related genes in three forms of ovine paratuberculosis using real-time PCR analyses and confirm that sheep pauci- and multibacillary disease are linked to type 1 and type 2 T cell responses respectively. The expression patterns of other cytokines shows that both disease forms have an inflammatory aetiology but that the central role for IL-1 alpha in bovine paratuberculosis is not seen in the sheep infection. Asymptomatic animals are infected and show no pathology but can be distinguished, in terms of cytokine expression pattern, from uninfected controls.

Figure 5: Statistically significant changes in genes between the three IS900+ groups and the uninfected control group. (a) Comparison of paucibacillary and control; (b) comparison of multibacillary and control; (c) comparison of asymptomatic and control. Results are given as significant (p ≤ 0.05) fold-changes of mean copy-numbers relative to the mean copy-numbers of the comparative group.

Mentions:
This is more clearly shown in Figures 4 and 5 where the results are expressed as statistically significant (p = 0.05) fold change where data are compared in six pairs – paucibacillary vs asymptomatic, multibacillary vs asymptomatic, paucibacillary vs multibacillary (Figure 4), paucibacillary vs control, multibacillary vs control and asymptomatic vs control (Figure 5). The expression levels of two genes, IL-1α and GM-CSF was relatively consistent in all the animals regardless of disease status.

Figure 5: Statistically significant changes in genes between the three IS900+ groups and the uninfected control group. (a) Comparison of paucibacillary and control; (b) comparison of multibacillary and control; (c) comparison of asymptomatic and control. Results are given as significant (p ≤ 0.05) fold-changes of mean copy-numbers relative to the mean copy-numbers of the comparative group.

Mentions:
This is more clearly shown in Figures 4 and 5 where the results are expressed as statistically significant (p = 0.05) fold change where data are compared in six pairs – paucibacillary vs asymptomatic, multibacillary vs asymptomatic, paucibacillary vs multibacillary (Figure 4), paucibacillary vs control, multibacillary vs control and asymptomatic vs control (Figure 5). The expression levels of two genes, IL-1α and GM-CSF was relatively consistent in all the animals regardless of disease status.

Bottom Line:
No changes were seen in IL-1 alpha in any sheep ileum tissues.Asymptomatic animals are IS900+ with normal histology but have significantly decreased levels of IL-18 and increased levels TNFalpha.Asymptomatic animals are infected and show no pathology but can be distinguished, in terms of cytokine expression pattern, from uninfected controls.

Background: Johne's disease is a chronic inflammatory disease of the gut caused by infection with Mycobacterium avium subspecies paratuberculosis (MAP). Symptoms include wasting, diarrhoea, loss of condition and eventual death. Three forms of Johne's disease have been described in sheep - paucibacillary, multibacillary and asymptomatic. The paucibacillary form is characterized by an inflammatory, Th1-type immune response. The multibacillary form of the disease, which disseminates the infection, is characterized by macrophage infiltration mediated by a Th2-type immune response, and asymptomatic animals have no clinical symptoms or pathology but are infected with MAP. What determines these three forms of the disease is unknown. To further understand these differences, we used real-time RT-PCR to compare the expression of thirteen cytokine and cytokine-related genes in ileal tissue from sheep with the three forms of the disease.

Results: Three pathological forms of sheep paratuberculosis were defined on the basis of histopathology, cytochemistry (Zeihl-Neelsen) and IS900 PCR. Paucibacillary lesions have largely T cell and eosinophil infiltration and are ZN negative; multibacillary lesions have macrophage infiltration and large numbers of acid-fast bacteria. The pauci- and multibacillary forms are linked to the differential expression of IFN gamma and IL-10 respectively. In addition the increased levels of the proinflammatory cytokines (IL-1 beta and TNFalpha), IL-8, IL-18 and TRAF-1 in both diseased forms is indicative of persistent inflammatory lesions. No changes were seen in IL-1 alpha in any sheep ileum tissues. Asymptomatic animals are IS900+ with normal histology but have significantly decreased levels of IL-18 and increased levels TNFalpha.

Conclusion: We have quantified the expression levels of thirteen cytokine and cytokine related genes in three forms of ovine paratuberculosis using real-time PCR analyses and confirm that sheep pauci- and multibacillary disease are linked to type 1 and type 2 T cell responses respectively. The expression patterns of other cytokines shows that both disease forms have an inflammatory aetiology but that the central role for IL-1 alpha in bovine paratuberculosis is not seen in the sheep infection. Asymptomatic animals are infected and show no pathology but can be distinguished, in terms of cytokine expression pattern, from uninfected controls.